This Program Project Grant application is founded on the hypothesis that the injury and recovery of the alveolar epithelial cell barrier are of crucial importance in determining the outcome of patients with the acute respiratory distress syndrome (ARDS). In these patients, different regions of the alveolar epithelium are exposed to hypoxia, hyperoxia, stretch and mechanical shear, all of which may contribute to alveolar epithelial cell injury. We have assembled a highly interactive group of senior investigators to study alveolar epithelial cell injury in these models via five interrelated projects. In Project # 1 we will determine the mechanisms by which hypoxia inhibits the Na,K-ATPase via phosphorylation-endocytosis and ubiquitination-degradation of the Na+ pump resulting in impaired lung liquid clearance. In Project # 2 we will determine the mechanisms by which exposure of alveolar epithelial cells to cyclic stretch or cyclic shear stress causes injury by altering the structure and function of intermediate filaments, the major protein comprising the cell cytoskeleton. In Project # 3 we will determine the mechanisms by which laminins and integrins at the cell surface serve as mechanosignaling molecules transducing MAPK signals in alveolar epithelial cells during cyclic stretch. In Project #4 we will investigate the role of receptor-dependent and mitochondrial-dependent alveolar epithelial apoptosis in the development of lung injury following exposure to hyperoxia. In Project # 5 we will determine the mechanisms by which exposure of epithelial cells to cyclic stretch enhances gene transfer to the alveolar epithelium. Collaborative studies have been conducted for each of the projects and the preliminary results support the feasibility of this proposal. This Program Project focuses the multidisciplinary expertise of the assembled investigators on the elucidation of mechanisms contributing to lung epithelial cell injury in clinically relevant models. These projects are interactive conceptually and pragmatically, where the aggregate of the projects is greater than the sum of its parts. The collective data will provide a composite picture of the regulation of the major alveolar epithelial cell proteins (the Na,K-ATPase and intermediate filaments represent about 60% of total alveolar epithelial cell proteins), determine the role of epithelial apoptosis in the pathogenesis of lung injury, develop novel strategies for the non-viral delivery of genes to the lung and define the role of the extracellular matrix in mechanosignaling in the alveolar epithelium. The insights gained from the data generated from these studies will provide novel molecular targets for the development of new therapeutic strategies to treat patients with lung injury.